Figure S3.
Altered cell cycle timing and anaphase spindle orientation after release from a prolonged metaphase arrest.(A) Representative time-lapse images showing an unperturbed control cell released from α factor. Numbers indicate time in minutes, with time 0 indicating the time of spindle breakdown in the first division cycle. Scale bar, 5 µm. (B) Representative time-lapse images showing a cell that was first released from α factor, then underwent a 3-h metaphase arrest, and then was released from the metaphase arrest. In this cell, the anaphase spindle elongates from the mother into the bud. Time 0 represents the time of anaphase spindle breakdown after metaphase release. Scale bar, 5 µm. (C) The timing of Cdc42 polarization, bud formation, and anaphase onset measured from the first spindle breakdown (time 0) after α factor release of unperturbed cells (n = 55 cells from two experiments). Error bars indicate SD. ****, P < 0.0001; **, P = 0.006 (Mann-Whitney t test). (D) The timing of Cdc42 polarization, bud formation, and anaphase onset measured from spindle breakdown (time 0) after the release from a 3-h metaphase arrest. Cells have MET3prCDC20, which allows a metaphase block with Cdc20 depletion in methionine-containing medium. The release occurs when methionine is removed from the medium. Error bars indicate SD (n = 55 cells from two experiments). ****, P < 0.0001 (Mann-Whitney t test). ns, not statistically significant. (E) The timing of Cdc42 polarization, bud formation, and anaphase onset measured from the first spindle breakdown (time 0) after α factor release (unperturbed) or metaphase release (metaphase released) daughter cells. Error bars indicate SD (n = 55 cells from two experiments). ****, P < 0.0001; ***, P = 0.002; *, P < 0.0273 (Mann-Whitney t test). (F) The timing of Cdc42 polarization, bud formation, and anaphase onset measured from the first spindle breakdown (time 0) after α factor release (unperturbed) or metaphase release (metaphase released) mother cells. Error bars indicate SD (n = 55 cells from two experiments). ****, P < 0.0001; ns, not statistically significant (Mann-Whitney t test). (G) Orientation of anaphase spindle elongation in unperturbed cells and cells released from a 3-h metaphase arrest (n = 55 cells from two experiments).

Altered cell cycle timing and anaphase spindle orientation after release from a prolonged metaphase arrest. (A) Representative time-lapse images showing an unperturbed control cell released from α factor. Numbers indicate time in minutes, with time 0 indicating the time of spindle breakdown in the first division cycle. Scale bar, 5 µm. (B) Representative time-lapse images showing a cell that was first released from α factor, then underwent a 3-h metaphase arrest, and then was released from the metaphase arrest. In this cell, the anaphase spindle elongates from the mother into the bud. Time 0 represents the time of anaphase spindle breakdown after metaphase release. Scale bar, 5 µm. (C) The timing of Cdc42 polarization, bud formation, and anaphase onset measured from the first spindle breakdown (time 0) after α factor release of unperturbed cells (n = 55 cells from two experiments). Error bars indicate SD. ****, P < 0.0001; **, P = 0.006 (Mann-Whitney t test). (D) The timing of Cdc42 polarization, bud formation, and anaphase onset measured from spindle breakdown (time 0) after the release from a 3-h metaphase arrest. Cells have MET3prCDC20, which allows a metaphase block with Cdc20 depletion in methionine-containing medium. The release occurs when methionine is removed from the medium. Error bars indicate SD (n = 55 cells from two experiments). ****, P < 0.0001 (Mann-Whitney t test). ns, not statistically significant. (E) The timing of Cdc42 polarization, bud formation, and anaphase onset measured from the first spindle breakdown (time 0) after α factor release (unperturbed) or metaphase release (metaphase released) daughter cells. Error bars indicate SD (n = 55 cells from two experiments). ****, P < 0.0001; ***, P = 0.002; *, P < 0.0273 (Mann-Whitney t test). (F) The timing of Cdc42 polarization, bud formation, and anaphase onset measured from the first spindle breakdown (time 0) after α factor release (unperturbed) or metaphase release (metaphase released) mother cells. Error bars indicate SD (n = 55 cells from two experiments). ****, P < 0.0001; ns, not statistically significant (Mann-Whitney t test). (G) Orientation of anaphase spindle elongation in unperturbed cells and cells released from a 3-h metaphase arrest (n = 55 cells from two experiments).

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